Atherosclerosis is a disease of the blood vessel wall that causes heart attacks, strokes, and loss of limbs. Despite major advances in medical and surgical management, atherosclerosis still causes significant morbidity and mortality. The impact of atherosclerosis on the health of the American people is expected to increase in the coming decades due to aging of the population and the consequences of the current obesity/diabetes epidemic. The broad, long-term objective of this project is to develop a novel therapy that decreases the morbidity and mortality of atherosclerosis. This novel therapy involves the introduction and expression of disease-preventing genes in the cells that line the blood vessel wall, and is appropriately termed atheroprotective gene therapy. Blood vessels treated with atheroprotective gene therapy would not develop atherosclerosis because they have been genetically modified to resist the underlying biological processes that cause atherosclerosis, including inflammation and accumulation of fat deposits in the blood vessel wall. There are 3 specific aims, all of which are carried out in rabbits. The aims are focused on developing clinically useful atheroprotective gene therapy, delivered by a promising gene-transfer vector, helper-dependent adenovirus (HDAd). HDAd is an attractive vector for human gene therapy because it expresses therapeutic genes stably for years in animals (including nonhuman primates) and is relatively non-inflammatory. The 3 specific aims exploit the promise of HDAd to address critical issues in preclinical gene-therapy research: 1) demonstration of long-term efficacy and lack of toxicity in large animal models of human disease (Aims 1 and 3); and 2) achievement of transgene expression levels that are sufficiently high to treat and prevent chronic diseases while minimizing the need for infusion of large amount of vectors (Aim 2). The first specific aim uses 2 animal models of robust atherosclerotic lesion growth and a model of atherosclerosis regression to test whether expression of apolipoprotein A-I in arterial endothelial cells can prevent growth of new atherosclerotic lesions and promote regression of existing lesions. The second specific aim develops improved expression cassettes that allow HDAd to express therapeutic genes at higher, more stable levels, specifically in endothelial cells. The third specific aim tests whether HDAd can achieve persistent expression of an atheroprotective gene in the clinically relevant setting of venous bypass grafting. Accomplishment of these 3 aims will bring clinical vascular gene therapy closer to implementation. Moreover, achievement of the aims will establish new experimental animal models and novel vector platforms for expressing therapeutic genes. These new models and vectors will facilitate development-both by our laboratory and others-of gene-based therapies that prevent and reverse human vascular disease.